A vast body of literature indicates a clear role for CD4+ T cells in the pathology of demyelinating disease. Many have investigated functional responses of these T cells in response to various antigens of the central nervous system (CNS) but the affinity these cells have for myelin remains understudied. Considering T cell affinity for antigen is correlated with the magnitude of the immune response, many have attempted to analyze this affinity with major histocompatibility complex (MHC) class II tetramers and other assays. Recently these methods have been shown to be inefficient at detecting functionally responsive CD4+ T cells of low antigen affinity in a polyclonal T cell population. In the proposed study, we will investigate the role of CD4+ T cell frequency and affinity in a secondary progressive demyelinating disease course. The non-obese diabetic (NOD) mouse model is clinically relevant to autoimmune demyelinating disease in its MHC-class II-associated proclivity to autoimmune disease and its tendency to follow a secondary progressive disease course that resembles that seen in the majority of human multiple sclerosis (MS) patients. Using the NOD model, we will quantify the frequency and affinity of CD4+ T cells for CNS antigens throughout the course of symptom relapse, remission and chronic progression. To accomplish this we will use the novel micropipette frequency adhesion assay which provides a sensitive and physiologically relevant two dimensional (2D) measurement of T-cell receptor affinity for peptide:MHC in a cell-anchored system. By investigating the role low-affinity CD4+ T cells play in the NOD model we hope to better understand the mechanism behind symptom relapse and remission observed in secondary progressive demyelinating disease. We have outlined a strong training plan that together with the accomplishment of the research aims prepares the fellowship applicant to attain her career goals of being a research-oriented clinical neurologist with a focus on immunological disease and inflammation in the CNS particularly the immunology and treatment of MS.